{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,13]],"date-time":"2026-08-13T13:37:30Z","timestamp":1786628250756,"version":"build-2736575974"},"reference-count":28,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2026,6,3]],"date-time":"2026-06-03T00:00:00Z","timestamp":1780444800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2026,8,13]],"date-time":"2026-08-13T00:00:00Z","timestamp":1786579200000},"content-version":"vor","delay-in-days":71,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>\n                    Infestations of\n                    <jats:italic>Cochliomyia hominivorax<\/jats:italic>\n                    , the New World Screwworm (NWS), has become a reemerging veterinary concern in the United States due to the recent northward expansion of fly detections as far as northern Mexico. Rapid, accurate and validated detection pipelines need to be developed in the case of an incursion into the United States. Confirmatory cases are evaluated by morphological identification with no paired test to verify identifications. With the frequency of submissions of non-ideal samples, particularly from fly traps, a molecular tool would be necessary for species identification. In this manuscript, we develop and assess a pipeline including three real-time PCR assays targeting the ribosomal RNA and five sets of Sanger primers targeting the mitochondrial genome that would be used as a paired tool with morphological identification. Two of the assessed real-time PCR assays are highly specific, sensitive and repeatable requiring\u2009&lt;\u20091 copy per reaction for detection. Four of the five Sanger primer sets were assessed, optimized and results evaluated for potential use in preliminary geographic analysis of specimens. This workflow will expedite screening of samples, provide a method to verify results using different tools and help understand genetic variations within the mitochondria for NWS outbreaks.\n                  <\/jats:p>","DOI":"10.1038\/s41598-026-54866-6","type":"journal-article","created":{"date-parts":[[2026,6,3]],"date-time":"2026-06-03T10:56:10Z","timestamp":1780484170000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Validation of a molecular workflow for Cochliomyia hominivorax (New World screwworm) identification in field samples"],"prefix":"10.1038","volume":"16","author":[{"given":"Rachel","family":"Palinski","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jessica","family":"Hicks","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jeffery T.","family":"Alfred","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alec","family":"Thompson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Patrick","family":"Camp","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julia","family":"Thomas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gleeson","family":"Murphy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Suelee","family":"Robbe-Austerman","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,6,3]]},"reference":[{"key":"54866_CR1","unstructured":"New world screwworm (Cochliomyia hominivorax). WOAH - World Organisation for Animal Health https:\/\/www.woah.org\/en\/disease\/new-world-screwworm-cochliomyia-hominivorax\/"},{"key":"54866_CR2","unstructured":"CDC. Clinical Overview of New World Screwworm. New World Screwworm (2025). https:\/\/www.cdc.gov\/new-world-screwworm\/hcp\/clinical-overview\/index.html"},{"key":"54866_CR3","unstructured":"World Organisation for Animal Health (WOAH). New world screwworm (cochliomyia hominivorax) and old world screwworm (chrysomya bezziana). Chap. 3.1.15. in Terrestrial animal health code (Paris (France, 2022)."},{"key":"54866_CR4","unstructured":"Harris, B. lastz\/lastz. (2026)."},{"key":"54866_CR5","doi-asserted-by":"crossref","unstructured":"Song, H., Buhay, J. E., Whiting, M. F. & Crandall, K. A. Many species in one: DNA barcoding overestimates the number of species when nuclear mitochondrial pseudogenes are coamplified. Proc. Natl. Acad. Sci. U. S. A. 105, 13486\u201313491 (2008).","DOI":"10.1073\/pnas.0803076105"},{"key":"54866_CR6","unstructured":"Wu, S. & Manber, U. AGREP-A fast approximate pattern-matching tool. (1992). https:\/\/www.usenix.org\/legacy\/publications\/library\/proceedings\/wu.pdf"},{"key":"54866_CR7","doi-asserted-by":"publisher","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","volume":"25","author":"H Li","year":"2009","unstructured":"Li, H. et al. The Sequence Alignment\/Map format and SAMtools. Bioinformatics 25, 2078\u20132079 (2009).","journal-title":"Bioinformatics"},{"key":"54866_CR8","unstructured":"R Core Team. R A Language and Environment for Statistical Computing. R Foundation for Statistical Computing, Vienna. - References - Scientific Research Publishing. (2021). https:\/\/www.scirp.org\/reference\/referencespapers?referenceid=3131254"},{"key":"54866_CR9","doi-asserted-by":"crossref","unstructured":"Hickner, P. V. et al. Haplotype-resolved genome assemblies for the New World screwworm, Cochliomyia hominivorax (Diptera: Calliphoridae), using the trio binning approach. G3 GenesGenomesGenetics 16, jkag053 (2026).","DOI":"10.1093\/g3journal\/jkag053"},{"key":"54866_CR10","doi-asserted-by":"publisher","first-page":"1345","DOI":"10.1016\/S0020-7519(02)00095-4","volume":"32","author":"D Otranto","year":"2002","unstructured":"Otranto, D. & Stevens, J. R. Molecular approaches to the study of myiasis-causing larvae. Int. J. Parasitol. 32, 1345\u20131360 (2002).","journal-title":"Int. J. Parasitol."},{"key":"54866_CR11","unstructured":"Screwworm.gov | Unified Government Response To Protect the United States. https:\/\/www.aphis.usda.gov\/livestock-poultry-disease\/stop-screwworm"},{"key":"54866_CR12","doi-asserted-by":"publisher","first-page":"23819","DOI":"10.1038\/s41598-025-04804-9","volume":"15","author":"UM Valdez-Espinoza","year":"2025","unstructured":"Valdez-Espinoza, U. M. et al. The reemergence of the New World screwworm and its potential distribution in North America. Sci. Rep. 15, 23819 (2025).","journal-title":"Sci. Rep."},{"key":"54866_CR13","doi-asserted-by":"publisher","first-page":"89","DOI":"10.1079\/BER2001135","volume":"92","author":"SR Skoda","year":"2002","unstructured":"Skoda, S. R., Pornkulwat, S. & Foster, J. E. Random amplified polymorphic DNA markers for discriminating Cochliomyia hominivorax from C. macellaria (Diptera: Calliphoridae). Bull. Entomol. Res. 92, 89\u201396 (2002).","journal-title":"Bull. Entomol. Res."},{"key":"54866_CR14","doi-asserted-by":"publisher","first-page":"772","DOI":"10.1139\/g98-164","volume":"42","author":"ME Infante-Malachias","year":"1999","unstructured":"Infante-Malachias, M. E. & Yotoko, K. S. C. Lima De Azeredo Espin, A. M. Random amplified polymorphic DNA of screwworm fly populations (Diptera: Calliphoridae) from Southeastern Brazil and Northern Argentina. Genome 42, 772\u2013779 (1999).","journal-title":"Genome"},{"key":"54866_CR15","doi-asserted-by":"publisher","unstructured":"Haymer, D. S. Random Amplified Polymorphic Dnas and Microsatellites: What Are They, and Can They Tell Us Anything We Don\u2019t Already Know? (2025). https:\/\/doi.org\/10.1093\/aesa\/87.6.717","DOI":"10.1093\/aesa\/87.6.717"},{"key":"54866_CR16","unstructured":"Cochliomyia hominivorax Probe qPCR Kit. Biofargo https:\/\/biofargo.com\/products\/skygen-cochliomyia-hominivorax-probe-qpcr-kit-without-internal-control"},{"key":"54866_CR17","unstructured":"Christen, J. A. Molecular-based identification of the New World screwworm, Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae). ETD Collect. Univ. Neb. -Linc. 1\u2013224 (2008)."},{"key":"54866_CR18","unstructured":"Driver, L. et al. A novel real-time PCR for New World screwworm fly (Cochliomyia hominivorax) and its application in a non-destructive multiplex for efficient detection of screwworm flies. Med. 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